Proper diagnosis of urinary tract infections depends on obtaining an uncontaminated urine sample for culture. Suprapubic aspiration and transurethral catheterization are the 2 recommended procedures for obtaining specimens for urine culture from young infants. The objective of the current study was to compare the pain that is experienced during these 2 procedures when performed in young infants. A prospective, single-blind, randomized, controlled study was conducted at a university-affiliated hospital in Israel. Institutional Research Ethics Board approved the study. Infants who were 0 to 2 months of age and presented to the emergency department with fever and therefore required urine collection for culture were randomly assigned evenly into 2 sample collection groups: suprapubic aspiration or transurethral catheterization. Patients were excluded when they were born prematurely or had had a previous sepsis workup or other painful procedures or an anomaly of the urogenital system or abdominal wall. Eutectic mixture of local anesthetic cream that contained lidocaine and prilocaine was applied 1 hour before the procedure. The urethra was catheterized using a 5-Fr latex-free feeding tube that was lubricated with sterile water-soluble jelly that contained 2% lidocaine hydrochloride. Pediatric residents who were experienced with the procedures performed both suprapubic aspiration and transurethral catheterization. The parents were instructed to use any comfort strategies that they wished, including verbal or physical comforting and pacifiers. Pain during collection was assessed on a 100-mm visual analog scale by a nurse and a parent. In addition, the infant's upper part of the body was videotaped during the procedure. An investigator, who was blinded to the procedure, assigned a point score according to the Douleur Aigue du Nouveaune neonatal acute pain scale. For ensuring a successful blinding process, the following steps were taken. First, camera recording started 30 seconds before the procedure to prevent the possibility of distinguishing between the procedures on the basis of their duration. Second, the physician and the nurse were asked not to speak during the procedure to avoid revealing the nature of the procedure. Third, the person who videotaped the procedure watched the tape before it was analyzed to ensure the impossibility of identifying the procedure from the tape. The Student's t test was used to compare the groups. The primary outcome was the mean Douleur Aigue du Nouveaune score. Secondary outcomes were the mean visual analogue scale for pain as estimated by the parents and by the nurse. We estimated that 25 patients would be needed in each group to detect a difference in the mean Douleur Aigue du Nouveaune score of at least 2 points with a power of 80% and alpha of .05. The study was conducted between April 1, 2004, and April 30, 2005. Fifty-eight infants were recruited; 29 were randomly assigned to suprapubic aspiration, and 29 were randomly assigned to transurethral catheterization. Seven infants were excluded because of consent withdrawal (3 patients), because of technical difficulties during videotaping (3 patients), or because the child voided during the procedure (1 patient). Twenty-seven infants in the suprapubic aspiration group and 24 in the transurethral catheterization group completed the study. All male infants were circumcised. An adequate urine sample was obtained in 18 (66%) of 27 patients in the suprapubic aspiration group and in 20 (83.3%) of 24 in the transurethral catheterization group. The mean Douleur Aigue du Nouveaune score was significantly higher in patients who were randomly assigned to suprapubic aspiration compared with patients who were randomly assigned to transurethral catheterization (7 and 4.5, respectively). The differences in Douleur Aigue du Nouveaune score also were significant in a subgroup analysis of boys and girls. Mean visual analogue scale scores by parents was higher in the suprapubic aspiration group compared with transurethral catheterization (63 +/- 27 mm vs 46 +/- 26, respectively). Similarly, mean visual analogue scale scores by nurses was higher in the suprapubic aspiration group compared with transurethral catheterization (3 +/- 18 mm vs 43 +/- 25 mm, respectively). In infants who are younger 2 months, suprapubic aspiration is more painful than transurethral catheterization. Health professionals should consider these differences when choosing a method for obtaining a urine sample from young infants.